title: Alpha-tocopherol supplementation of macrophages does not influence their ability to oxidize LDL.
abstract: We have investigated the effect of alpha-tocopherol-loading of mouse peritoneal macrophages and human monocytes on their ability to oxidize human low density lipoprotein (LDL). Mouse peritoneal macrophages incorporated alpha-tocopherol (alpha-TOH) from culture medium supplemented with the vitamin in a time- and concentration-dependent manner. Subcellular fractionation by density gradient ultracentrifugation showed that the distribution of incorporated alpha-TOH within the cell was similar to that of free cholesterol. Most (approximately 88%) of alpha-TOH partitioned into the membrane fractions (plasma membrane approximately 41%, mitochondria and lysosomes approximately 26%, and endosomes plus endoplasmic reticulum approximately 21%). Cellular alpha-TOH was stable for at least 24 h in serum- or LDL-free media whether permissive (Ham's F-10) or non-permissive (Dulbecco's minimum essential medium, DMEM) for LDL oxidation. When incubated with LDL in DMEM, alpha-TOH-preloaded cells transferred small amounts of alpha-TOH (approximately 1 nmol/mg LDL protein after 9 h) to the lipoprotein. However, enrichment of the cells with alpha-TOH did not change the kinetics of oxidation of either normal or TOH-depleted LDL in Ham's F-10 medium compared with non-loaded cells, as assessed by alpha-TOH consumption, cholesteryl ester degradation, and cholesteryl ester hydroperoxide and 7-ketocholesterol accumulation. Nor did it alter superoxide release by the cells or their ability to reduce extracellular copper(II). Similar to mouse macrophages, enrichment of human monocytes with alpha-TOH did not change the kinetics of cell-mediated LDL oxidation. We conclude that elevated cellular levels of alpha-TOH in mouse peritoneal macrophages and in human monocytes do not affect their ability to oxidize LDL lipids in vitro. This suggests that either cell-mediated oxidation of LDL under the conditions of this study is not dependent on cell-derived radical species or that cellular alpha-TOH is unable to affect their formation.
authors.affiliation: Cell Biology Unit, The Heart Research Institute, Camperdown, New South Wales, Australia.
authors.forename: A
authors.initials: A
authors.lastname: Baoutina
authors.forename: R T
authors.initials: RT
authors.lastname: Dean
authors.forename: W
authors.initials: W
authors.lastname: Jessup
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D003300:Copper / Q000378:metabolism; D003470:Culture Media; D004721:Endoplasmic Reticulum / Q000378:metabolism; D011992:Endosomes / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008077:Lipoproteins, LDL / Q000378:metabolism*; D008084:Lipoxygenase / Q000378:metabolism; D008247:Lysosomes / Q000378:metabolism; D008264:Macrophages / Q000187:drug effects* / Q000378:metabolism* / Q000648:ultrastructure; D017737:Macrophages, Peritoneal / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D008928:Mitochondria / Q000378:metabolism; D009000:Monocytes / Q000187:drug effects / Q000378:metabolism; D010084:Oxidation-Reduction; D013481:Superoxides / Q000378:metabolism; D014810:Vitamin E / Q000378:metabolism / Q000494:pharmacology*
chemical_list: D003470:Culture Media; D008077:Lipoproteins, LDL; C099252:oxidized low density lipoprotein; D013481:Superoxides; D014810:Vitamin E; D003300:Copper; D008084:Lipoxygenase